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Innovative strategies to control malaria are urgently needed. Exploring the interplay between Plasmodium sp. parasites and host red blood cells (RBCs) offers opportunities for novel antimalarial interventions. Pyruvate kinase deficiency (PKD), characterized by heightened 2,3-diphosphoglycerate (2,3-DPG) concentration, has been associated with protection against malaria. Elevated levels of 2,3-DPG, a specific mammalian metabolite, may hinder glycolysis, prompting us to hypothesize its potential contribution to PKD-mediated protection. We investigated the impact of the extracellular supplementation of 2,3-DPG on the Plasmodium falciparum intraerythrocytic developmental cycle in vitro. The results showed an inhibition of parasite growth, resulting from significantly fewer progeny from 2,3-DPG-treated parasites. We analyzed differential gene expression and the transcriptomic profile of P. falciparum trophozoites, from in vitro cultures subjected or not subjected to the action of 2,3-DPG, using Nanopore Sequencing Technology. The presence of 2,3-DPG in the culture medium was associated with the significant differential expression of 71 genes, mostly associated with the GO terms nucleic acid binding, transcription or monoatomic anion channel. Further, several genes related to cell cycle control were downregulated in treated parasites. These findings suggest that the presence of this RBC-specific glycolytic metabolite impacts the expression of genes transcribed during the parasite trophozoite stage and the number of merozoites released from individual schizonts, which supports the potential role of 2,3-DPG in the mechanism of protection against malaria by PKD.

The Mycobacterium abscessus complex (MABC) is an emerging, difficult to treat, multidrug-resistant nontuberculous mycobacteria responsible for a wide spectrum of infections and associated with an increasing number of cases worldwide. Dominant circulating clones (DCCs) of MABC have been genetically identified as groups of strains associated with higher prevalence, higher levels of antimicrobial resistance, and worse clinical outcomes. To date, little is known about the genomic characteristics of MABC species circulating in Portugal. Here, we examined the genetic diversity and antimicrobial resistance profiles of 30 MABC strains isolated between 2014 and 2022 in Portugal. The genetic diversity of circulating MABC strains was assessed through a gene-by-gene approach (wgMLST), allowing their subspecies differentiation and the classification of isolates into DCCs. Antimicrobial resistance profiles were defined using phenotypic, molecular, and genomic approaches. The majority of isolates were resistant to at least two antimicrobials, although a poor correlation between phenotype and genotype data was observed. Portuguese genomes were highly diverse, and data suggest the existence of MABC lineages with potential international circulation or cross-border transmission. This study highlights the genetic diversity and antimicrobial resistance profile of circulating MABC isolates in Portugal while representing the first step towards the implementation of a genomic-based surveillance system for MABC at the Portuguese NIH.

Guava, pitanga and acerola are known for their vitamin content and high levels of bioactive compounds. Thus, the preparation of combinations of these fruits comprises a blend with high nutraceutical potential, yielding a strong and attractive pigmentation material. In this study, the influence of different proportions of maltodextrin on the lyophilization of a blend of guava, acerola and pitanga was evaluated considering not only the physicochemical, physical and colorimetric parameters but also the bioactive compounds in the obtained powders. The blend was formulated from the mixture and homogenization of the three pulps in a ratio of 1:1:1 (m/m), then maltodextrin was added to the blend, resulting in four formulations: blend without adjuvant (BL0), and the others containing 10% (BL10), 20% (BL20) and 30% (BL30) maltodextrin. The formulations were lyophilized and disintegrated to obtain powders. The powders were characterized in terms of water content, water activity, pH, total titratable acidity, ash, total and reducing sugars, ascorbic acid, total phenolic content, flavonoids, anthocyanins, carotenoids, lycopene, color parameters, Hausner factor, Carr index, angle of repose, solubility, wettability and porosity. All evaluated powders showed high levels of bioactive compounds and the increase in maltodextrin concentration promoted positive effects, such as reductions in water content, water activity and porosity and improved flow, cohesiveness and solubility characteristics.

The combination of fruit pulps from different species, in addition to multiplying the offer of flavors, aromas and textures, favors the nutritional spectrum and the diversity of bioactive principles. The objective was to evaluate and compare the physicochemical characteristics, bioactive compounds, profile of phenolic compounds and in vitro antioxidant activity of pulps of three species of tropical red fruits (acerola, guava and pitanga) and of the blend produced from the combination. The pulps showed significant values of bioactive compounds, with emphasis on acerola, which had the highest levels in all parameters, except for lycopene, with the highest content in pitanga pulp. Nineteen phenolic compounds were identified, being phenolic acids, flavanols, anthocyanin and stilbene; of these, eighteen were quantified in acerola, nine in guava, twelve in pitanga and fourteen in the blend. The blend combined positive characteristics conferred by the individual pulps, with low pH favorable for conservation, high levels of total soluble solids and sugars, greater diversity of phenolic compounds and antioxidant activity close to that of acerola pulp. Pearson's correlation between antioxidant activity and ascorbic acid content, total phenolic compounds, flavonoids, anthocyanins and carotenoids for the samples were positive, indicating their use as a source of bioactive compounds.

Among the vegetables that stand out for their high concentration of anthocyanins, red cabbage appears as one of the most-used sources of these pigments in food production and it is considered a suitable raw material for the extraction of natural dye. Therefore, the objective was to carry out the production of natural extracts from red cabbage, under different conditions, varying the solvent, type of pre-treatment, pH range, and processing temperature during the concentration of the extracts. The anthocyanins were extracted from red cabbage using the following solvents: distilled water, 25% ethyl alcohol, and 70% ethyl alcohol. The raw material was divided into two groups, the first was subjected to a drying pre-treatment at 70 °C for 1 h and for the second group, the extraction was performed with the raw material in natura. Two pH ranges of 4.0 and 6.0 and extraction temperatures of 25 °C and 75 °C were used in the extracts, resulting in 24 formulations. The extracts obtained were analyzed for colorimetric parameters and anthocyanins. The results of anthocyanins show that the methodology that uses 25% alcohol, pH 4.0, and processing temperature of 25 °C produces a reddish extract and better results in the extraction, presenting average values of 191.37 mg/100 g of anthocyanins, being 74% higher compared to the highest values obtained in the other extracts where the same raw material was used and the solvents differed.

The application of whole genome sequencing of Mycobacterium tuberculosis directly on clinical samples has been investigated as a means to avoid the time-consuming need for culture isolation that can lead to a potential prolonged suboptimal antibiotic treatment. We aimed to provide a proof-of-concept regarding the application of the molecular capture of M. tuberculosis genomes directly from positive sputum samples as an approach for epidemiological and drug susceptibility predictions. Smear-positive sputum samples ( n = 100) were subjected to the SureSelectXT HS Target Enrichment protocol (Agilent Technologies, Santa Clara, CA, USA) and whole-genome sequencing analysis. A higher number of reads on target were obtained for higher smear grades samples (i.e., 3+ followed by 2+). Moreover, 37 out of 100 samples showed ≥90% of the reference genome covered with at least 10-fold depth of coverage (27, 9, and 1 samples were 3+, 2+, and 1+, respectively). Regarding drug-resistance/susceptibility prediction, for 42 samples, ≥90% of the >9000 hits that are surveyed by TB-profiler were detected. Our results demonstrated that M. tuberculosis genome capture and sequencing directly from clinical samples constitute a potential valid backup approach for phylogenetic inferences and resistance prediction, essentially in settings when culture is not routinely performed or for samples that fail to grow.

The study of human papillomavirus (HPV)-induced carcinogenesis uses multiple in vivo mouse models, one of which relies on the cytokeratin 14 gene promoter to drive the expression of all HPV early oncogenes. This study aimed to determine the HPV16 variant and sublineage present in the K14HPV16 mouse model. This information can be considered of great importance to further enhance this K14HPV16 model as an essential research tool and optimize its use for basic and translational studies. Our study evaluated HPV DNA from 17 samples isolated from 4 animals, both wild-type (n = 2) and HPV16-transgenic mice (n = 2). Total DNA was extracted from tissues and the detection of HPV16 was performed using a qPCR multiplex. HPV16-positive samples were subsequently whole-genome sequenced by next-generation sequencing techniques. The phylogenetic positioning clearly shows K14HPV16 samples clustering together in the sub-lineage A1 (NC001526.4). A comparative genome analysis of K14HPV16 samples revealed three mutations to the human papillomaviruses type 16 sublineage A1 representative strain. Knowledge of the HPV 16 variant is fundamental, and these findings will allow the rational use of this animal model to explore the role of the A1 sublineage in HPV-driven cancer.

Dissemination of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in healthcare institutions affects both patients and health-care workers (HCW), as well as the institutional capacity to provide essential health services. Here, we investigated an outbreak of SARS-CoV-2 in a "non-COVID-19" hospital ward unveiled by massive testing, which challenged the reconstruction of transmission chains. The contacts network during the 15-day period before the screening was investigated, and positive SARS-CoV-2 RNA samples were subjected to virus genome sequencing. Of the 245 tested individuals, 48 (21 patients and 27 HCWs) tested positive for SARS-CoV-2. HCWs were mostly asymptomatic, but the mortality among patients reached 57.1% (12/21). Phylogenetic reconstruction revealed that all cases were part of the same transmission chain. By combining contact tracing and genomic data, including analysis of emerging minor variants, we unveiled a scenario of silent SARS-CoV-2 dissemination, mostly driven by the close contact within the HCWs group and between HCWs and patients. This investigation triggered enhanced prevention and control measures, leading to more timely detection and containment of novel outbreaks. This study shows the benefit of combining genomic and epidemiological data for disclosing complex nosocomial outbreaks, and provides valuable data to prevent transmission of COVID-19 in healthcare facilities.

This work addresses the problem of tracking a signal-emitting mobile target in wireless sensor networks (WSNs) with navigated mobile sensors. The sensors are properly equipped to acquire received signal strength (RSS) and angle of arrival (AoA) measurements from the received signal, while the target transmit power is assumed not known. We start by showing how to linearize the highly non-linear measurement model. Then, by employing a Bayesian approach, we combine the linearized observation model with prior knowledge extracted from the state transition model. Based on the maximum a posteriori (MAP) principle and the Kalman filtering (KF) framework, we propose new MAP and KF algorithms, respectively. We also propose a simple and efficient mobile sensor navigation procedure, which allows us to further enhance the estimation accuracy of our algorithms with a reduced number of sensors. Model flaws, which result in imperfect knowledge about the path loss exponent (PLE) and the true mobile sensors' locations, are taken into consideration. We have carried out an extensive simulation study, and our results confirm the superiority of the proposed algorithms, as well as the effectiveness of the proposed navigation routine.

Leishmaniases are a group of neglected tropical diseases (NTDs) caused by protozoan parasites from >20 Leishmania species. Visceral leishmaniasis (VL), also known as kala-aza, is the most severe form of leishmaniasis, usually fatal in the absence of treatment in 95% of cases. The Morita-Baylis-Hillman adducts (MBHAs) are being explored as drug candidates against several diseases, one of them being leishmaniasis. We present here the design, synthesis and in vitro screening against Leishmania donovani of sixteen new molecular hybrids from analgesic/antiinflammatory tetrahydropyrans derivatives and Morita-Baylis-Hillman adducts. First, acrylates were synthesized from analgesic/anti-inflammatory tetrahydropyrans using acrylic acid under TsOH as a catalyst (70-75% yields). After the 16 new MBHAs were prepared in moderate to good yields (60-95%) promoted by microwave irradiation or low temperature (0 °C) in protic and aprotic medium. The hybrids were evaluated in vitro on the promastigote stage of Leishmania donovani by determining their inhibitory concentrations 50% (IC 50 ), 50% hemolysis concentration (HC 50 ), selectivity index (HC 50 /IC 50 ,), and comparing to Amphotericin B, chosen as the anti-leishmanial reference drug. The hybrid which presents the bromine atom in its chemical structure presents high leishmanicide activity and the high selectivity index in red blood cells (SI rb > 180.19), compared with the highly-toxic reference drug (SI rb = 33.05), indicating that the bromine hybrid is a promising compound for further biological studies.